SCL GENE AND HEMATOPOIETIC DEVELOPMENT

本研究将探讨基因的表达和功能， SCL基因在造血发育过程中的蛋白产物 在老鼠。 该基因首先在一个突变位点的断裂点上被发现。 人类T细胞白血病染色体易位，是 转录因子家族，其成员被认为是 分化的调节剂。 这些实验的结果应该 提供对造血的基本机制的深入了解。 以来 通过染色体重排激活该基因是最常见的 急性T细胞白血病的遗传性损害，这些研究也应该 有助于我们理解白血病的发生 第一特定 这项工作的目的是表征的翻译产品的 小鼠SCL基因在造血细胞中的表达。 的蛋白质产品 鼠SCL基因将在造血细胞系中表征， 小鼠组织通过放射免疫沉淀和Western印迹分析。 SCL蛋白的亚细胞定位将通过以下方法确定： COS细胞和稳定表达的细胞系中的瞬时表达 单个蛋白质。 第二个具体目标是描述 小鼠SCL基因的转录和翻译产物 在造血发育过程中。 将研究SCL mRNA表达 具有RNA酶保护、原位杂交和聚合酶链 免疫细胞化学方法检测SCL蛋白的表达。 小鼠胚胎分期 转录和翻译产物 也将在体外模型中表征基因的特征， 使用胚胎干（ES）细胞衍生的造血发育 胚状体 第三个具体目标是确定以下行动： SCL蛋白与造血发育 这将被调查 在ES细胞的功能获得和功能丧失突变体中， 进行分化。 第四个具体目标是确定 SCL蛋白作为转录调节因子的功能 在造血发育过程中。 SCL蛋白质的能力， 反式激活人工和生理靶点（如果确定）， 被确定。 某些SCL蛋白可能具有 不同的转录能力，并作为转录 将研究拮抗剂。 细胞目标将被识别 使用基于分离SCL-诱导的功能策略， 启动子，以及不同SCL同种型激活这种启动子的能力。 发起人将被定义。

The proposed research will investigate the expression and functions of the protein products of the SCL gene during hematopoietic development in the mouse. This gene was first identified at the breakpoint of a chromosomal translocation in human T-cell leukemia and is a member of a family of transcription factors whose members are recognized to be regulators of differentiation. The results of these experiments should provide insight into basic mechanisms of hematopoiesis. Since activation of this gene by chromosomal rearrangement is the most common genetic lesion in acute T-cell leukemia, these studies should also contribute to our understanding of leukemogenesis. The first specific aim of this work is to characterize the translational products of the murine SCL gene in hematopoietic cells. The protein products of the murine SCL gene will be characterized in hematopoietic cell lines and mouse tissues by radioimmunoprecipitation and Western blot analysis. Subcellular localization of SCL proteins will be determined by transient expression in COS cells and in cell lines stably expressing individual proteins. The second specific aim is to characterize the transcriptional and translational products of the murine SCL gene during hematopoietic development. SCL mRNA expression will be studied with RNase protection, in situ hybridization, and the polymerase chain reaction (PCR) and SCL protein expression by immunocytochemistry in staged mouse embryos. The transcriptional and translational products of the gene will also be characterized in an in vitro model of hematopoietic development using embryonic stem (ES) cell-derived embryoid bodies. The third specific aim is to define the actions of SCL proteins in hematopoietic development. This will be investigated in gain of function and loss of function mutants of ES cells made to undergo differentiation. The fourth specific aim is to determine the ability of SCL proteins to function as transcriptional regulators during hematopoietic development. The ability of SCL proteins to transactivate artificial and, if identified, physiologic targets will be determined. The possibility that certain SCL proteins may have different transcriptional potencies and act as transcriptional antagonists will be investigated. Cellular targets will be identified using a functional strategy based on the isolation of SCL-inducible promoters, and the ability of different SCL isoforms to activate such promoters will be defined.